This project aims to clarify further the cellular mechanisms of epileptogenesis in mammalian neocortex in vivo. For this purpose, a newly devised method for topical application of penicillin that results in predictable extracellular penicillin concentrations will be used to produce small, discrete, epileptogenic foci. With this technique, penicillin gradients of known magnitude are produced across the soma and apical dendritic arborizations of pyramidal cells. In the experiments proposed here, we will characterize physiologically identified neurons in terms of active and passive membrane electrical properties including input resistance, impedance, rheobase current, spike-initiation threshold, charging time constant, and accommodation, and determine the effect of penicillin on these measurements. Using a mathematical model of the diffusional kinetics of penicillin in neocortex, we will characterize changes in membrane biophysics in terms of local penicillin concentration. Patterns of cell discharge during focal epileptogenesis will be correlated with particular cell types in morphologic studies using intracellular horseradish peroxidase (HRP). By correlating the developing spatial distribution of different forms of paroxysmal depolarization shifts (PDS) with the concentration distributions of penicillin during evolving foci, we will test our hypothesis that the local penicillin concentration is a better indicator of the type of interictal activity generated by a neuron than its location within the neocortex. Finally, we will test the hypothesis that penicillin initiates PDSs in neocortical pyramidal cells in vivo mainly through loss of inhibition by antagonism of GABA-mediated IPSPs at or near the soma. This will be done by recording from pyramidal cells in the presence of a penicillin gradient where apical dendritic arborizations are at effective (30mM, layer I) concentrations for GABA antagonism while somata are at ineffective (2mM or less, layers V-VI) concentration. These recordings will be compared to those done in a reversed gradient formed by diffusion of penicillin from a deep source deposited by microinjection. Iontophoresis of GABA, GABA agonists, and antagonists will help delineate this mechanism. HRP will be injected to verify cell type and to locate the cell.